2, 4-diamino-6-amido-s-triazines



United States Patent (Hi 3,350,398 2,4-DIAMINO-6-AMlD0-s-TRIAZINES Takeo Ueda, 1066 Izumi, Komae-cho, Kitatama-gun; Shigeshi Toyoshima, 355 Funabashi-cho, Setagaya-ku; and Mitsuru Furukawa, 897 Yukigaya-cho, Ota-ku; all of Tokyo, Japan N Drawing. Original application Mar. 28, 1963, Ser. No. 268,551, new Patent No. 3,271,393, dated Sept. 6, 1966. and this application Mar. 3, 1966, Ser. No. 9 Claims priority, application Japan, July 3, 1962, 37/27,619 Claims. (Cl. 260-2493) This invention relates to new chemical compounds which are useful in inhibiting and/or inactivating measles, echo and coxsackie viruses. More particularly, this invention relates to 2,4-diaminoand 2-amino-4-(substituted)- amino-6- N-carb onyl -s-triazine.

This application is a division of application Serial No. 268,551, filed March 28, 1963, now US. Patent No. 3,271,393.

It is an object of this invention to provide a new class of chemical compounds which possess antiviral activity, and more particularly, compounds which inhibit and/or inactivate viruses which cause measles.

There are provided, according to the present invention, compounds having the formula wherein R and R are members selected from the group taken together may be a heterocyclic group containing from 4 to 5 carbon atoms; and Y is a member selected from the group consisting of -OH,

in which R and R are members selected from the group consisting of morpholino, piperidino, pyrrolidino, hydrogen, alkyl, alkenyl, aralkyl and aryl. The group taken together may be, for example,

morpholino (O N) plperidino N- and pyrrolidlno 3,350,398 Patented Oct. 31, 1967 ice Formula I is -NHCNH,

is and the group taken together is morpholino (i.e.,

The compounds of the present invention (except those wherein Y is -OH) may be prepared by the reaction of an int-substituted biguanide having the formula II R1 NH NH NiiNH-ii-NH= R2 wherein R and R are the same as defined above, with a di( lower)alkyl oxalate, such as diethyl oxalate. The

III R NH solvent an a-substituted biguanide having the Formula II above, a di(lower)alkyl oxalate and ammonia or an amine selected from the group consisting of NH R in which R and R are the same as defined above.

The compounds represented by Formula II are known in the art. For example, when R and R are both hydrogen, the compound is biguanide which is commercially available. When the group taken together is morpholino (i.e.,

which is the preferred embodiment of this invention, the compound is 4-morpholinecarboximidoylguanidine which is described in Chemical Abstracts, 54, 10131 and 21510.

Another method for making the novel compounds of this invention (except those wherein Y is -OH) involves the reaction of a compound having the formula in which R and R are the same as defined above and X is a member selected from the group consisting of chloro, bromo, fluoro, iodo, and -OR (R being alkyl), with ammonia or an amine selected from the group consisting of R3 NH C--NH: and EN NH R in which R and R pounds of Formula tained by the reaction of a compound of Formula III with an alcohol; or by the reaction of a compound of Formula II, a di(lower)alky1 oxalate and an alcohol.

Compounds represented by Formula I wherein Y is hydroxyl may be synthesized by the hydrolysis of a compound of Formula IV in which X is -OR or by the hydrolysis of a compound of Formula III, or by reacting a compound of Formula II with an oxalic acid monoester. The product from any of these reactions may be further reacted with a halogenation reagent to obtain a compound of Formula IV in which X is halogen.

The compounds of this invention possess useful antiviral activity, particularly virucidal activity. The compounds of the present invention are effective in inhibiting the multiplication of the measles virus (Edmonstan strain). They have also been found to be effective against echo and coxsackie viruses. The compounds of this invention are also useful in the study of viruses, particularly as a tool to facilitate study of their basic biological characteristics, including their intracellular multiplication.

The following examples will illustrate the preparation of the compounds of the present invention.

EXAMPLE 1 A solution of sodium methoxide (0.1 mole) in absolute methanol is added, with heating and stirring, to a suspension of 0.1 mole of powdered 4-morpholinecarboximidoylguanidine hydrochloride in absolute methanol. NaCl which is formed as a precipitate is removed by filtration. Diethyl oxalate (0.1 mole) is then added and the solution is refluxed for 30 minutes on a steam bath. The reaction which occurs may be represented by the following equation:

The reaction mixture is allowed to stand overnight and the precipitated product is collected by filtration. The product is found to have a melting point of 268 C., a yield of 79%.

Elemental analysis.-Calculated for C H O N N, 31.10%. Found, 31.02%.

A suspension of 0.01 mole of the product obtained above is refluxed in 150 cc. of absolute ethanol until all of the material has been dissolved. T o the solution there is added 0.01 mole of guanidine and the mixture is refluxed for 5 hours. A white precipitate is formed during the refluxing. The reaction which occurs may be represented by the following equation:

The reaction mixture is concentrated, the precipitate recovered and recrystallized from water. The product, 2- amino-4-morpholino-6 (N-amidinocarbamoyl) -s-triazine, consists of prisms having a melting point of 271 C.

Elemental analysis.-Calculated for C H O N 42.08%. Found: N, 41.50%.

The inhibitory effect of 2-amino-4-morpholino-6-(N- amidinocarbamoyl)-s-triazine on the multiplication of polio virus (type 1Mahoney and type 2-MEF and measles (Edmonstan strain) is determined using the standard tissue culture method. KB cells were used for polio and Hep. N0. 2 cells for measles. Table I sets forth the Tissue Culture Inhibitory Dose (TCID) for the compound tested as well as for the control.

TABLE I TCID50/0J 1. Viral Material Control Treated Polio:

Type 1-Mahoney 10 10- Type 2MEF1 10- 10- Measles: Edmonstan 10- 10- In another test procedure, the minimum inhibitory concentration of 2-amino-4 morph0lino-6 (N-amidinocarbamoyl)-s-triazine on measles virus in Hep. No. 2 cells is determined. The results are set forth in Table II.

(In the results column, the denominator denotes the total number of assay tubes used in the experiment and the numerator denotes the number of tubes in which no cytopathogenic effect was observed.)

The minimum inhibitory concentration of 2-amino-4- morpholino-6 (N-amidinocarbamoyl) s-triazine on the Malhoney strain of virus (10- is found to be 4x10- mo e.

The following examples are illustrative of the preparation of various intermediates of the type represented for Formula III, above:

EXAMPLE 2 Preparation of 1,1-dimethyl-3-(4,5-di0x0-2- imidazolidinylidene)-guanidine Sodium ethoxide prepared from 0.23 g. of metallic sodrum and a small amount of dehydrated ethanol is added, with warming, to a suspension of 1.65 g. of 1,1-dimethyl 6 biguanide hydrochloride in cc. of dehydrated ethanol. EXAMPLE 9 After the precipitated NaCl is filtered off, 1.46 g. of dig precipitate is col suction. Yield 1.13 g. (62%), M.P. 266267 decomposition).

Analysis.Calcd. N. 38.28. Found: C,

EXAMPLE 3 Preparation of .I,]-(2,2-0xydiethyl) Z-imidazolidinylidene) -3-(4,5-dioxo- -guanidine To a solution of 21 g. of 1,l-(2,2-oxydiethyl) bi'guanide hydrochloride 1n 1 l. of dehydrated methanol is oxide prepared from 2.5 g. of metal le volume of dehydrated methanol. Cl is removed by filtration. After addition of g. of diethyl oxalate to the filtrate and allowing to stand for 2 hours, the resulting precipitate is collected by suction; yield 19.8 g. (79%), M.P. 268 C. (with decomposition).

Analysis.Calcd. for C H O N N, 31.10. Found:

EXAMPLE 4 Preplaration of l-(p-m'ethoxyphenyl) sodium and a suita The precipitated Na tion mixture is collected by filtration; yield 2.40 g. (92% M.P. 208 C. (with decomposition).

Analysis.Calcd. for C H O N N, 26.81. Found: N, 27.05.

Other examples of intermediates of the type represented by Formula HI which are prepared by the general method described above are set forth in Table III.

Preparation of 4-amino-6-isopropylamin0-striazine-Z-carbanilide A suspension of 1.97 g. of 1 imidazolidinylidene)-guanidine i on a water bath until completel pletion of the reacti -isopropyl-3-(4,5-dioxo-2- n 5 cc. of aniline is heated y dissolved. After a com- Preparation of 4-amino-6-cyclohexylamino-s-triazine-Z darbOxypiperidide O -N H (5 A mixture of 2.3 imidazolidinylidene) heated for 1 hour 0 tate is collected by from ethanol to col yield 1.64. g. (54%) Analyszlsu-Calcd. for C H ON N, 27.61. Found: N, 27.89.

7 g. of 1-cyclohexyl-3-(4,5-dioxo-2- -guanidine and 1 cc. of piperidine is n a water bath. The resulting precipisuction on cooling and recrystallized orless prisms which melt at 266 C EXAMPLE 1 1 Preparation of 4-amin0-6- (p amino) -s-triazine TABLE III R Analysis N, percent Group N Melting Yield Molecular Example Point (percent) Formula R2 C.) Cale. Exper. of Formula 1115 a NE2- 300 C4H502N5 45.15 45.50 0 iso-CdEbNH- 21s 11 C7HuO2N'5 35.52 35.00 1 @NH 249 as CZOH102N 29.52 29.80 s one-Grim- 214 ourruoim 28.56 28.86

The following examples are illustrative of the preparation of various compounds represented by Formula I, above:

p-methoxyphenyl) -3- 4,5 -guanidine and 1 cc. of pyrr on a water bath. After cooling, the deposited precipitate is collected by filtration and recrystallized from ethanol to colorless needles which melt at 215-216 C.; yield 1.82 g. (58%).

Analysis.-Calcd. for C 5H O N N, 26.74. Found: N, 26.73.

EXAMPLE 12 Preparation of 4-amino-6-(N-rnorpholinyl)-striazine-Z-carboxyguanidide hydrate EXAMPLE 13 Preparation of 4-amin0-6-(p-t0luidin0)- s-triazine-2-carboxypyrrolidide A mixture of 2.45 g. of l-(p-tolyl) -3-(4,5-dioxo-2- imidazolidinylidene)-guanidine and 0.71 g. of pyrrolidine is heated for 2 hours on a water bath. After a completion of the reaction, the resulting residue is recrystallized from a small amount of ethanol to give colorless prisms which melt at 258 C.; yield 1.24 g. (50%).

Analysis.-Calcd. for C H ON N, 28.34.

N, 28.17. Found:

EXAMPLE 14 Preparation of 4-am'in0-6- p-toluidino) s-triazine-Z-carboxylic acid N iL-NH:

N CHa-QNH- Procedure 1.-2.45 g. irnidazolidinylidene)-guauidine is dissolved in a alkaline aqueous solution and the solution is neutralized with hydrochloric acid. The resulting precipitate is collected by suction, washed with hot water and dried at C. under pressure; M.P. 234-235 C. (with decomposition) Analysis.-Calcd. for C H O N C, 53.87; H, 4.52; N, 28.56. Found: C, 53.21; H, 4.29; N, 28.48.

Procedure 2.2.7 g. of ethyl 4-amino-6-(p-toluidino)- s-triazine-2-carboxylate is refluxed with ethanolic sodium hydroxide solution for 5 hours. After cooling the precipitate is collected and dissolved in a small amount of water, followed by neutralization with hydrochloric acid, and the colorless crystals precipitated are collected by suction, M.P. 234-235" C.

Procedure 3.-A mixture of 1.91 g. of 1-(p-tolyl)- biguanide and 1.51 g. of potassium salt of monoethyl oxalate in dehydrated ethanol is refluxed for 10 hours. When cool, the precipitate is filtered and dissolved in a small amount of water. The solution is neutralized with hydrochloric acid and the colorless crystals precipitated are collected by suction, M.P. 234235 C.

EXAMPLE 1 5 (Procedure 1) Preparation of ethyl 4-amin0-6-(N- morpholinyl)-s-triazine-Z-carboxylate 2.25 g. of 1,l-(2,2-oxydiethyl)-3-(4,5-dioxo-2-imidazolidinylidene)guanidine in cc. of dehydrated ethanol is refluxed for 10 hours. The mixture is concentrated, followed by chilling. The precipitate is filtered and recrystallized from ethanol to obtain colorless prisms; yield 1.62 g., M.P. 172 C.

Analysis.-Calcd. for C H O N N, 27.66. Found: N, 28.13.

(Procedure 2) Preparation of 4-amin0-6-(N- morpholinyl)-s-triazine-2-carboxyguanidide To a solution of 2.53 g. of ethyl 4-amino-6-(N-morpholinyl)-s-triazine-2-carboxylate in 100 cc. of dehydrated ethanol, 0.6 g. of guanidine was added. After refluxing on a water bath for 5 hours, the mixture was concentrated and chilled. The precipitate was collected by suction, recrystallized from water to colorless prisms; M.P. 271 C.

EXAMPLE 16 Preparation of 4-amin0-6-(p-t0luidin0) -s-triazine-2- carboxypyrrolidide To 2.45 g. of 4-amino-6-(p-toluidino) -s-triazine-2- carboxylic acid prepared by the above method is added an excess amount of thionyl chloride. After removal of excess thionyl chloride in vacuo, 1.5 cc. of pyrrolidine is added to the residue, and warmed on a water bath for 2 hours. After cooling, a diluted solution of alkali is added to the mixture, the resulting precipitate is recrystallized from ethanol to colorless prisms; yield 1.8 g., M.P. 258 C.

Further examples of compounds of this invention represented by Formula I and which are prepared by the general methods described above are set forth in dilute 75 Table IV.

3,350,398 11 12 Z-amino-4-N-n-butylamin0-6-(N-amidinocar- 2. The compound of claim 1 having the formula bamoyl) -s-triazine, O Z-amino-4-isobutylamino-6- (N-amidinocarbamoyl) I s-triazine, 2-amino-4- (N-pentyl-N-t-butylamino)-6- (N- amidinocarbamoyl)striazine 3Hs 2-am1no-fl-hexylammo-6-(N-arnidinocarbamoyl)- IJJ NH2 s-trlazine, 2-amino-4-(Z-ethylhexylamino)-6-(N-amidinocar- CH: N

'bamoyl)-s-triazine, 1 3. The compound of claim 1 having the formula 2-amino-4-piperidino-6- (N-amidinocarbamoyl) s-triazine, 2-amino-4-pyrrolidino-6- (N-amidinocarbamoyl) H s-triazine, and 2-amino-4-cyclohexylamino-6-(N-amidinocar- We claim:

1. A compound selected from the group consisting of The compound of claim 1 having an formula compounds of the formula (I) W C E N N Y b CH OQNH-l i l-NH,

R1 N N N a g 5. The compound of claim 1 having the formula /N -NH: o W \N LNG? N% N wherein R and R are members selected from the group J: H consisting of hydrogen, (lower) alkyl, phenyl, (lower) Q alkyl-substituted phenyl, (lower) alkoxy-substituted phenyl and cyclohexyl, and Y is a member selected from the group consisting of morpholino, piperidino, pyrrolidino, References Clted (lowefhlkylamino, Phenylamino, and a d cal of the 40 Smolin'eta .,s-Triazines and Derivatives, Interseience formula Pub. Inc., New York, (1959), pp. 244-252.

WALTER A. MODANCE, Primary Examiner. F J. D. RANDOLPH, Examiner. J. M. FORD, Assistant Examiner. 

1. A COMPOUND SELECTED FROM THE GROUP CONSISTING OF COMPOUNDS OF THE FORMULA 